We have used asymmetric quantum dots (QD) to conduct imaging measurements of individual 2H3 cell Type I Fcε receptor rotation on timescales down to 10 ms per frame and have used time-tagged single photon counting measurements of individual QD to examine μs timescales, although such rapid timescales are limited by QD emission rates. In both approaches, decays of time-autocorrelation functions (TACF) for fluorescence polarization fluctuations extend into the millisecond timescale, as implied by time-resolved phosphorescence anisotropy results. Results obtained following crosslinking of the Type I Fcε receptor suggest that this slow decay may be a property of the membrane itself, perhaps reflecting large-scale fluctuations of mesoscale membrane regions. However, depending on instrumental parameters used in data analysis, polarization fluctuation TACFs can contain a contribution from the intensity fluctuation TACF arising from QD blinking. QD blinking feed-through is extremely sensitive to these analysis parameters which effectively change slightly from one measurement to another. We discuss approaches based on the necessary statistical independence of polarization and intensity fluctuations to guarantee removal of a blinking-based component from rotation measurements. Supported by NSF grant MCB-1024668.

Blinking of quantum dot probes in measurements of molecular rotation on cell surfaces

249th National Meeting (2015)

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